Corallococcus caeni sp. nov., a novel myxobacterium isolated from activated sludge.

Two myxobacterial strains (KH5-1T and NO1) were isolated from the activated sludge tanks treating municipal sewage wastewater in Japan. These strains were recognised as myxobacteria based on their phenotypic characteristics of swarming colonies and fruiting bodies. Phylogenetic analyses using the 16S rRNA gene revealed that strains KH5-1T and NO1 were affiliated with the genus Corallococcus, with the closest neighbours being Corallococcus exercitus AB043AT (99.77% and 99.84%, respectively). Genome comparisons using orthologous average nucleotide identity (orthoANI) and digital DNA-DNA hybridisation similarity (dDDH) with strains KH5-1T and NO1 and their phylogenetically close relatives in Corallococcus spp. were below the thresholds. The major cellular fatty acids of strains KH5-1T and NO1 were iso-C15:0 (31.9%, 30.0%), summed feature 3 (comprising C16:1ω7c and/or C16:1ω6c) (20.2%, 17.7%), and iso-C17:0 (12.1%, 14.8%), and the major respiratory quinone was found to be menaquinone (MK)-8. Based on the phenotypic, chemotaxonomic, and phylogenetic evidence, strains KH5-1T and NO1 represent a new species in the genus Corallococcus, for which the proposed name is Corallococcus caeni sp. nov. The type strain is KH5-1T (= NCIMB 15510T = JCM 36609T).

Corallococcus senghenyddensis sp. nov., a myxobacterium with potent antimicrobial activity.

AIM: Corallococcus species are diverse in the natural environment with 10 new Corallococcus species having been characterized in just the last 5 years. As well as being an abundant myxobacterial genus, they produce several secondary metabolites, including Corallopyronin, Corramycin, Coralmycin, and Corallorazine. We isolated a novel strain Corallococcus spp RDP092CA from soil in South Wales, UK, using Candida albicans as prey bait and characterized its predatory activities against pathogenic bacteria and yeast. METHODS AND RESULTS: The size of the RDP092CA genome was 8.5 Mb with a G + C content of 71.4%. Phylogenetically, RDP092CA is closely related to Corallococcus interemptor, C. coralloides, and C. exiguus. However, genome average nucleotide identity and digital DNA-DNA hybridization values are lower than 95% and 70% when compared to those type strains, implying that it belongs to a novel species. The RDP092CA genome harbours seven types of biosynthetic gene clusters (BGCs) and 152 predicted antimicrobial peptides. In predation assays, RDP092CA showed good predatory activity against Escherichia coli, Pseudomonas aeruginosa, Citrobacter freundii, and Staphylococcus aureus but not against Enterococcus faecalis. It also showed good antibiofilm activity against all five bacteria in biofilm assays. Antifungal activity against eight Candida spp. was variable, with particularly good activity against Meyerozyma guillermondii DSM 6381. Antimicrobial peptide RDP092CA_120 exhibited potent antibiofilm activity with >50% inhibition and >60% dispersion of biofilms at concentrations down to 1 µg/ml. CONCLUSIONS: We propose that strain RDP092CA represents a novel species with promising antimicrobial activities, Corallococcus senghenyddensis sp. nov. (=NBRC 116490T =CCOS 2109T), based on morphological, biochemical, and genomic features.

Expanding the Myxochelin Natural Product Family by Nicotinic Acid Containing Congeners.

Myxobacteria represent a viable source of chemically diverse and biologically active secondary metabolites. The myxochelins are a well-studied family of catecholate-type siderophores produced by various myxobacterial strains. Here, we report the discovery, isolation, and structure elucidation of three new myxochelins N1-N3 from the terrestrial myxobacterium Corallococcus sp. MCy9049, featuring an unusual nicotinic acid moiety. Precursor-directed biosynthesis (PDB) experiments and total synthesis were performed in order to confirm structures, improve access to pure compounds for bioactivity testing, and to devise a biosynthesis proposal. The combined evaluation of metabolome and genome data covering myxobacteria supports the notion that the new myxochelin congeners reported here are in fact frequent side products of the known myxochelin A biosynthetic pathway in myxobacteria.

Diazotrophic Anaeromyxobacter Isolates from Soils.

Biological nitrogen fixation is an essential reaction in a major pathway for supplying nitrogen to terrestrial environments. Previous culture-independent analyses based on soil DNA/RNA/protein sequencing could globally detect the nitrogenase genes/proteins of Anaeromyxobacter (in the class Deltaproteobacteria), commonly distributed in soil environments and predominant in paddy soils; this suggests the importance of Anaeromyxobacter in nitrogen fixation in soil environments. However, direct experimental evidence is lacking; there has been no research on the genetic background and ability of Anaeromyxobacter to fix nitrogen. Therefore, we verified the diazotrophy of Anaeromyxobacter based on both genomic and culture-dependent analyses using Anaeromyxobacter sp. strains PSR-1 and Red267 isolated from soils. Based on the comparison of nif gene clusters, strains PSR-1 and Red267 as well as strains Fw109-5, K, and diazotrophic Geobacter and Pelobacter in the class Deltaproteobacteria contain the minimum set of genes for nitrogenase (nifBHDKEN). These results imply that Anaeromyxobacter species have the ability to fix nitrogen. In fact, Anaeromyxobacter PSR-1 and Red267 exhibited N2-dependent growth and acetylene reduction activity (ARA) in vitro Transcriptional activity of the nif gene was also detected when both strains were cultured with N2 gas as a sole nitrogen source, indicating that Anaeromyxobacter can fix and assimilate N2 gas by nitrogenase. In addition, PSR-1- or Red267-inoculated soil showed ARA activity and the growth of the inoculated strains on the basis of RNA-based analysis, demonstrating that Anaeromyxobacter can fix nitrogen in the paddy soil environment. Our study provides novel insights into the pivotal environmental function, i.e., nitrogen fixation, of Anaeromyxobacter, which is a common soil bacterium.IMPORTANCEAnaeromyxobacter is globally distributed in soil environments, especially predominant in paddy soils. Current studies based on environmental DNA/RNA analyses frequently detect gene fragments encoding nitrogenase of Anaeromyxobacter from various soil environments. Although the importance of Anaeromyxobacter as a diazotroph in nature has been suggested by culture-independent studies, there has been no solid evidence and validation from genomic and culture-based analyses that Anaeromyxobacter fixes nitrogen. This study demonstrates that Anaeromyxobacter harboring nitrogenase genes exhibits diazotrophic ability; moreover, N2-dependent growth was demonstrated in vitro and in the soil environment. Our findings indicate that nitrogen fixation is important for Anaeromyxobacter to survive under nitrogen-deficient environments and provide a novel insight into the environmental function of Anaeromyxobacter, which is a common bacterium in soils.

Simulacricoccus ruber gen. nov., sp. nov., a microaerotolerant, non-fruiting, myxospore-forming soil myxobacterium and emended description of the family Myxococcaceae.

A non-fruiting group of myxobacteria was previously speculated to exist in nature based on metagenomics data containing uncultured members of the order Myxococcales. Here, we describe a myxobacterial strain, designated MCy10636T, which was isolated from a German soil sample collected in 2013. It exhibits swarming characteristics but atypically produces myxospores in the absence of fruiting bodies. The novel strain stains Gram-negative and Congo-red-negative and is characterized mesophilic, neutrophilic, chemoheterotrophic and microaerotolerant. Branched-chain fatty acids are the predominant cellular fatty acids over the straight-chain type, and contain the major fatty acids iso-C17 : 0 2-OH, C16 : 1, iso-C17 : 0 and iso-C15 : 0. Based on blastn results, the 16S rRNA gene sequence reveals similarity (97 %) to Aggregicoccus edonensis MCy1366T, (97 %) Myxococcus macrosporus DSM 14697T, (96 %) Corallococcus coralloides DSM2259T and Corallococcus exiguus Cc e167T. Phylogenetic analysis showed a novel lineage of MCy10636T in the family Myxococcaceae, suborder Cystobacterineae. Based on polyphasic taxonomic characterization, we propose that this unusual, non-fruiting, myxospore-forming and microaerotolerant myxobacterial strain, MCy10636T, represents a novel genus and species, Simulacricoccus ruber gen. nov., sp. nov. (DSM 106554T=NCCB 100651T).

Nannocystis konarekensis sp. nov., a novel myxobacterium from an Iranian desert.

An orange-coloured myxobacterium, MNa11734T, was isolated from desert in Iran. MNa11734T had rod-shaped vegetative cells, moved by gliding and was bacteriolytic. No real fruiting body formation could be observed, but sporangioles were produced on water agar. The strain was mesophilic, strictly aerobic and chemoheterotrophic. 16S rRNA gene analyses revealed that MNa11734T belonged to the family Nannocystaceae, genus Nannocystis and was closely related to Nannocystis pusilla Na p29T (DSM 14622T) and Nannocystis exedens Na e1T (DSM 71T), with 97.8 and 97.6 % 16S rRNA gene sequence similarity, respectively. Laboratory-measured DNA-DNA hybridization showed only 9.5/15.7 % (reciprocal) similarity between the novel strain and N. pusilla Na p29T, and 14.1/20.4 % between the strain and N. exedens Na e1T, whereas DNA-DNA hybridization estimates derived from draft genome sequences were 21.8-23.0 % and 22.2-23.7 %, respectively, depending on the calculation method. The G+C content of DNA from Nannocystis konarekensis MNa11734T was 73.3 mol%, for N. pusilla Nap29T it was 71.8 mol% and for N. exedens Nae1T it was 72.2 mol%. The major fatty acids of the new strain were C16 : 1 (56.2 %), iso-C17 : 0 (14.4 %), C14 : 0 (8.2 %), C16 : 0 (6.6 %) and iso-C15 : 0 (5.9 %). Strain MNa11734T exhibited phylogenetic and physiological similarities to the two other species of Nannocystis, i.e. N. pusilla and N. exedens, but the differences were sufficient enough to represent a novel species, for which the name Nannocystiskonarekensis sp. nov. is proposed. The type strain is MNa11734T (=DSM 104509T=NCCB 100618T).

Actinobacteria and Myxobacteria-Two of the Most Important Bacterial Resources for Novel Antibiotics.

Bacteria have been by far the most promising resource for antibiotics in the past decades and will in all undoubtedly remain an important resource of innovative bioactive natural products in the future. Actinobacteria have been screened for many years, whereas the Myxobacteria have been underestimated in the past. Even though Actinobacteria belong to the Gram-positive and Myxobacteria to the Gram-negative bacteria both groups have a number of similar characters, as they both have huge genomes with in some cases more than 10kB and a high GC content and they both can differentiate and have often cell cycles including the formation of spores. Actinobacteria have been used for the antibiotic research for many years, hence it is often discussed whether this resource has now been exhaustively exploited but most of the screening programs from pharmaceutical companies were basing on the cultivation mainly of members of the genus Streptomyces or Streptomyces like strains (e.g., some Saccharopolyspora, Amycolatopsis or Actinomadura species) by use of standard methods so that many of the so called "neglected" Actinobacteria were overlooked the whole time. The present review gives an overview on the state of the art regarding new bioactive compounds with a focus on the marine habitats. Furthermore, the evaluation of Myxobacteria in our ongoing search for novel anti-infectives is highlighted.

Vitiosangium cumulatum gen. nov., sp. nov. and Vitiosangium subalbum sp. nov., soil myxobacteria, and emended descriptions of the genera Archangium and Angiococcus, and of the family Cystobacteraceae.

Bacterial strains MCy10943T and MCy10944T were isolated in 2014 from dried Nepalese soil samples collected in 2013 from Phukot, Kalikot, Western Nepal, and Godawari, Lalitpur, Central Nepal. The novel organisms showed typical myxobacterial growth characteristics, which include swarming colony and fruiting body formation on solid surfaces, and a predatory ability to lyse micro-organisms. The strains were aerobic, mesophilic, chemoheterotrophic and showed resistance to various antibiotics. The major cellular fatty acids common to both organisms were C17 : 0 2-OH, iso-C15 : 0, C16 : 1 and iso-C17 : 0. The G+C content of the genomic DNA was 72-75 mol%. Phylogenetic analysis showed that the strains belong to the family Cystobacteraceae, suborder Cystobacterineae, order Myxococcales. The 16S rRNA gene sequences of both strains showed 97-98 % similarity to Archangium gephyra DSM 2261T andCystobacter violaceus DSM 14727T, and 96.7-97 % to Cystobacter fuscus DSM 2262T and Angiococcus disciformis DSM 52716T. Polyphasic taxonomic characterization suggested that strains MCy10943T and MCy10944T represent two distinct species of a new genus, for which the names Vitiosangium cumulatum gen. nov., sp. nov. and Vitiosangium subalbum sp. nov. are proposed. The type strain of Vitiosangium cumulatum is MCy10943T (=DSM 102952T=NCCB 100600T) while that for Vitiosangium subalbum is MCy10944T (=DSM 102953T=NCCB 100601T). In addition, emended descriptions of the genera Archangium and Angiococcus, and of the family Cystobacteraceaeare provided.

Racemicystis persica sp. nov., a myxobacterium from soil.

A novel myxobacterium, strain MSr11462T, was isolated in 2015 from a soil sample collected form Kish Island beach, Persian Gulf, Iran. It displayed general myxobacterial features like Gram-negative staining, rod-shaped vegetative cells, gliding on solid surfaces, microbial lytic activity, fruiting-body-like aggregates and myxospore-like structures. The strain was mesophilic, aerobic and showed a chemoheterotrophic mode of nutrition. It was resistant to many antibiotics like gentamycin, polymyxin, fusidic acid and trimethoprim, and the key fatty acids of whole-cell hydrolysates were iso-C15 : 0, C16 : 0, iso-C17 : 0, C18 : 1, iso-C17 : 1 2-OH, C18 : 1 2-OH, iso-C15 : 0 OAG (O-alkylglycerol) and C16 : 1 OAG. The 16S rRNA gene sequence showed highest similarity (98.6 %) to Racemicystis crocea strain MSr9521T (GenBank accession no. KT591707). The phylogenetic analysis based on 16S rRNA gene sequences and matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) spectroscopy data supports a novel species of the family Polyangiaceae and the genus Racemicystis. DNA-DNA hybridization showed only about 50 % similarity between the novel strain and the phylogenetically closest species, Racemicystis. crocea MSr9521T. On the basis of a comprehensive taxonomic study, we propose a novel species, Racemicystis persica sp. nov., for strain MSr11462T (=DSM 103165T=NCCB 100606T).

Isolation and characterisation of the epothilone gene cluster with flanks from high alkalotolerant strain Sorangium cellulosum (So0157-2).

Epothilones are cytotoxic macrolactones having auspicious anti-tumorous activities, but merely produced by rare Sorangium strains. Here, we have focused on the epothilone gene cluster from special niche bacterial strain, S. cellulosum So0157-2. Therefore, we have isolated a high pH tolerant S. cellulosum strain So0157-2 and characterized the epothilones gene cluster and its flanks by cosmid/fosmid libraries preparation and sequencing. The assembly spanned 94,459 bp and consisted of 56,019 bp core region. Remarkably, the core as well as upstream 420 bp and downstream 315 bp were highly conserved, while further neighboring regions varied extremely. Transposase traces were identified near the core of clusters, supporting that the transposon-mediated transgenesis is a naturally evolved strategy for the cluster's dissemination. A predicted neighboring esterase gene was identified as a potential epothilone-resistance gene preventing self-toxicity. Novel modification or regulatory genes, a multi-position-cyclo releasing gene and their relationship with corresponding analogs were identified in strain So0157-2. These findings open the door to discover additional, naturally evolved epothilone-related genes for significant applications in industrial as well as clinical sector.

Racemicystis crocea gen. nov., sp. nov., a soil myxobacterium in the family Polyangiaceae.

A novel bacterial strain designated MSr9521T was isolated in 2014 from a soil sample collected in 1986 from the Philippines. The novel bacterium shows myxobacterial characteristics that include pseudoplasmodial swarming, fruiting body formation and predatory ability to lyse microorganisms. The strain is chemoheterotrophic, mesophilic and aerobic. Major fatty acids are C18:1, C17:1 2-OH and iso-C15:0, and also contains trace amounts of omega-3/-6 polyunsaturated fatty acids. The G+C content of the genomic DNA is 70.4 mol%. The 16S rRNA gene sequence shows 95-96 % closest similarity to Sorangium cellulosum DSM 14627T, Polyangium fumosum Pl fu5T, Jahnella thaxteri Pl t4T and Byssovorax cruenta By c2T. The molecular phylogenetic analysis shows that the novel isolate forms a novel branch in the family Polyangiaceae, suborder Sorangiineae. Polyphasic taxonomic characterization suggests that the strain MSr9521T represents a novel species of a new genus in the family Polyangiaceae, for which the name Racemicystis crocea gen. nov., sp. nov. is proposed. The type strain of Racemicystis crocea is MSr9521T (=DSM 100773T=NCCB 100574T).

Aggregicoccus edonensis gen. nov., sp. nov., an unusually aggregating myxobacterium isolated from a soil sample.

A novel myxobacterium, MCy1366(T) (Ar1733), was isolated in 1981 from a soil sample collected from a region near Tokyo, Japan. It displayed general myxobacterial features like Gram-negative-staining, rod-shaped vegetative cells, gliding on solid surfaces, microbial lytic activity, fruiting-body-like aggregates and myxospore-like structures. The strain was mesophilic, aerobic and showed a chemoheterotrophic mode of nutrition. It was resistant to many antibiotics such as cephalosporin C, kanamycin, gentamicin, hygromycin B, polymyxin and bacitracin, and the key fatty acids of whole cell hydrolysates were iso-C15 : 0, iso-C17 : 0 and iso-C17 : 0 2-OH. The genomic DNA G+C content of the novel strain was 65.6 mol%. The 16S rRNA gene sequence showed highest similarity (97.60 %) to 'Stigmatella koreensis' strain KYC-1019 (GenBank accession no. EF112185). Phylogenetic analysis based on 16S rRNA gene sequences and MALDI-TOF MS data revealed a novel branch in the family Myxococcaceae. DNA-DNA hybridization showed only 28 % relatedness between the novel strain and the closest recognized species, Corallococcus exiguus DSM 14696(T) (97 % 16S rRNA gene sequence similarity). A recent isolate from a soil sample collected in Switzerland, MCy10622, displayed 99.9 % 16S rRNA gene sequence similarity with strain MCy1366(T) and showed almost the same characteristics. Since some morphological features like fruiting-body-like aggregates were barely reproducible in the type strain, the newly isolated strain, MCy10622, was also intensively studied. On the basis of a comprehensive taxonomic study, we propose a novel genus and species, Aggregicoccus edonensis gen. nov., sp. nov., for strains MCy1366(T) and MCy10622. The type strain of the type species is MCy1366(T) ( = DSM 27872(T) = NCCB 100468(T)).

Minicystis rosea gen. nov., sp. nov., a polyunsaturated fatty acid-rich and steroid-producing soil myxobacterium.

A bacterial strain designated SBNa008(T) was isolated from a Philippine soil sample. It exhibited the general characteristics associated with myxobacteria, such as swarming of Gram-negative vegetative rod cells, fruiting body and myxospore formation and predatory behaviour in lysing micro-organisms. The novel strain was characterized as mesophilic, chemoheterotrophic and aerobic. The major fatty acids were C(20:4)ω6,9,12,15 all cis (arachidonic acid), iso-C(15 : 0), C(17 : 1) 2-OH and iso-C(15 : 0) dimethylacetal. Interestingly, SBNa008(T) contained diverse fatty acids belonging to the commercially valuable polyunsaturated omega-6 and omega-3 families, and a highly conjugated dihydroxylated C28 steroid. The G+C content of the genomic DNA was 67.3 mol%. The 16S rRNA gene sequence revealed 95-96% similarity to sequences derived from clones of uncultured bacteria and 94-95% similarity to cultured members of the suborder Sorangiineae. Phylogenetic analysis revealed that strain SBNa008(T) formed a novel lineage in the suborder Sorangiineae. Based on a polyphasic taxonomic characterization, we propose that strain SBNa008(T) represents a novel genus and species, Minicystis rosea gen. nov., sp. nov. The type strain of Minicystis rosea is SBNa008(T) ( =DSM 24000(T) =NCCB 100349(T)).

Vulgatibacter incomptus gen. nov., sp. nov. and Labilithrix luteola gen. nov., sp. nov., two myxobacteria isolated from soil in Yakushima Island, and the description of Vulgatibacteraceae fam. nov., Labilitrichaceae fam. nov. and Anaeromyxobacteraceae fam. nov.

Two myxobacterial strains (designated B00001(T) and B00002(T)) were isolated from forest soil samples collected from Yakushima Island, Kagoshima, Japan. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strains B00001(T) and B00002(T) respectively formed independent branches within the suborders Cystobacterineae and Sorangiineae and were most closely related to Cystobacter armeniaca DSM 14710(T) (90.4% similarity) and Byssovorax cruenta DSM 14553(T) (91.3%). Neither strain showed typical features of myxobacteria such as bacteriolytic action or fruiting body formation, but both had high DNA G+C contents (66.3-68.3 mol%). Swarming motility was observed in strain B00002(T) only. Cells of both strains were vegetative, chemoheterotrophic, mesophilic, strictly aerobic, Gram-negative, motile rods, and both strains exhibited esterase lipase (C8), leucine arylamidase, naphthol-AS-BI-phosphohydrolase and ß-galactosidase activities. Strain B00001(T) contained MK-7 as the predominant respiratory quinone and the major fatty acid was iso-C15:0. In contrast, strain B00002(T) contained MK-8 as the major cellular quinone and the major fatty acids were C16 : 1ω5c and iso-C17 : 0. Based on the phenotypic and genotypic data presented, strains B00001(T) and B00002(T) represent novel genera and species, for which we propose the names Vulgatibacter incomptus gen. nov., sp. nov. and Labilithrix luteola gen. nov., sp. nov., respectively. The type strains of Vulgatibacter incomptus and Labilithrix luteola are B00001(T) ( = NBRC 109945(T) = DSM 27710(T)) and B00002(T) ( = NBRC 109946(T) = DSM 27648(T)), respectively. The new genera are assigned to the new families Vulgatibacteraceae fam. nov. and Labilitrichaceae fam. nov., respectively. In addition, Anaeromyxobacteraceae fam. nov., is proposed to accommodate the genus Anaeromyxobacter, which is related to the genus Vulgatibacter.

[Isolation, identification and biological activity of myxobacteria from soils].

OBJECTIVE: Purpose of this work was to screen myxobacteria from soils and study their biological activities towards pathogenic bacteria, tumor cells and insects. METHODS: Through inactivated E. coli and filter paper inducing methods, we isolated and purified myxobacteria from soil samples. Then we identified these purified strains based on morphological observation, physiological and biochemical characteristics, and the 16S rDNA sequences homologous analysis. By plate diffusion experiments, oral toxicity tests and tetrazolium assays, we investigated the biological activities of the myxobacterial culture supernatant. RESULTS: We isolated 35 myxobacterial strains and classified them as 4 genera: Myxococcus (9), Corallococcus (9), Nannocystis (11) and Sorangium (6). Eight purified myxobacteria were identified and named. Cytotoxicity tests show that strain C. macrospores S22 had potent and broad-spectrum cytotoxic effect on tumor cell lines including B16, 4T1, HeLa and HCT-116, so did the strains M. fulvus S51, C. exiguus S22 and M. Xanthus S55. Additionally, C. macrospores S22 also shows inhibitory activity to pathogenic bacteria Bacillus subtillis and Candida albicans. CONCLUSION: Myxobacteria are widely distributed in natural soils. C. macrosporus has potent toxicity against cancer cells and pathogenic bacteria; and C. exiguus with antitumor activity. The myxobacterial strains are promising resources for discovery and development of new active natural products and drugs.

[Isolation and identification of Myxobacteria strain STXZ54 with antitumor activities].

OBJECTIVE: We isolated Myxobacteria strains from soil samples collected from Guangzhou, identified the strain and studied the antitumor activity. METHODS: We isolated Myxobacteria strains from soil samples through inactivated E. coli inducing method, identified the strain according to morphological observation, physiological and biochemical characteristics, and the homologous analysis of 16S rRNA sequences of nucleotides. The antitumor spectrum and the corresponding IC50 of the active component separated from the culture was analyzed. Confocal laser scanning microscope was used to examine the growth inhibitory effect of the active component on B16 cells. RESULTS: We isolated a Myxobacteria strain and identified as Myxococcus macrosporus STXZ54. The active component termed SGF5 showed cytostatic activity against B16, Hela, 4T1, Hep-3B with IC50 values of 10 microg/mL, and HCT-116 cell with IC50 values of 70 microg/mL. Subcellular structure of B16 cells were abnormal observed by confocal laser scanning microscope. Combining the apoptosis and necrosis assay result it is likely that SGF5 can induce apoptosis of B16 cells. CONCLUSION: The active component separated from the culture of Myxococcus macrosporus STXZ54 has a significant antitumor activity tested by cytotoxicity assay, which was worth exploiting as potential antitumor drugs.

Functional identification of rubber oxygenase (RoxA) in soil and marine myxobacteria.

The rubber oxygenase (RoxA) of Xanthomonas sp. strain 35Y (RoxA(Xsp)) is so far the only known extracellular c-type diheme cytochrome that is able to cleave poly(cis-1,4-isoprene). All other rubber-degrading bacteria described are Gram positive and employ a nonheme protein (latex-clearing protein [Lcp]) for the postulated primary attack of polyisoprene. Here, we identified RoxA orthologs in the genomes of Haliangium ochraceum, Myxococcus fulvus, Corallococcus coralloides, and Chondromyces apiculatus. The roxA orthologs of H. ochraceum (RoxA(Hoc)), C. coralloides BO35 (RoxA(Cco)), and M. fulvus (RoxA(Mfu)) were functionally expressed in a ΔroxA Xanthomonas sp. 35Y background. All RoxA orthologs oxidatively cleaved polyisoprene, as revealed by restoration of clearing-zone formation and detection of 12-oxo-4,8-dimethyltrideca-4,8-diene-1-al (ODTD) as a cleavage product. RoxA(Xsp), RoxA(Mfu), and RoxA(Cco) were purified and biochemically characterized. The optimal temperature of RoxA(Cco) and RoxA(Mfu) was between 22 and 30°C. All RoxA orthologs as isolated showed an oxidized UV-visible spectrum. Chemical reduction of RoxA(Cco) and RoxA(Mfu) indicated the presence of two slightly different heme centers with absorption maxima between 549 and 553 nm, similar to RoxA(Xsp). Sequence analysis and modeling of the three-dimensional structures of the RoxA orthologs revealed a high degree of similarity to the recently solved RoxA(Xsp) structure and included several conserved residues, notably, W302, F317, and a MauG motif at about H517. Lcp-like sequences were not detected in the genomes of the Xanthomonas sp. 35Y, H. ochraceum, M. fulvus, and C. coralloides. No RoxA orthologs were found in Gram-positive bacteria, and this first description of functional RoxA in Gram-negative bacteria other than Xanthomonas proves that RoxA is more common among rubber degraders than was previously assumed.

Pseudenhygromyxa salsuginis gen. nov., sp. nov., a myxobacterium isolated from an estuarine marsh.

A myxobacterial strain, designated SYR-2(T), was obtained from a mud sample from an estuarine marsh alongside the Yoshino River, Shikoku, Japan. It had rod-shaped vegetative cells and formed bacteriolytic enlarging colonies or so-called 'swarms' in the agar media. Fruiting-body-like globular to polyhedral cell aggregates and myxospore-like spherical to ellipsoidal cells within them were observed. Those features coincided with the general characteristics of myxobacteria. The strain was mesophilic and strictly aerobic. Growth of SYR-2(T) was observed at 18-40 °C (optimum, 30-35 °C), pH 5.5-8.3 (optimum, pH 7.0-7.5) and with 0.0-2.5 % (w/v) NaCl (optimum, 0.2-1.0 %). Both Mg(2+) and Ca(2+) were essential cations for the growth. The predominant fatty acids were iso-C15 : 0 (43.8 %), iso-C17 : 0 (22.4 %) and iso-C16 : 0 (9.6 %). A C20 : 4 fatty acid [arachidonic acid (4.3 %)], iso-C19 : 0 (1.5 %) and anteiso-acids [ai-C15 : 0 (0.5 %), ai-C17 : 0 (0.3 %)] were also detected. The G+C content of the DNA was 69.7 mol%. The strain contained menaquinone-7 (MK-7) as the major respiratory quinone. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain SYR-2(T) belonged to the suborder Nannocystineae, order Myxococcales in the class Deltaproteobacteria, and the strain was most closely related to two type strains of marine myxobacteria, Enhygromyxa salina SHK-1(T) and Plesiocystis pacifica SIR-1(T), with 96.5 % and 96.0 % similarities, respectively. These characteristics determined in this polyphasic study suggested that strain SYR-2(T) represents a novel species in a new genus of myxobacteria. The name Pseudenhygromyxa salsuginis gen. nov., sp. nov. is proposed to accommodate this isolate, and the type strain of Pseudenhygromyxa salsuginis is SYR-2(T) ( = NBRC 104351(T) = DSM 21377(T)).

[Influence of different prey strains on isolation myxobacteria in saline-alkaline soils of Xinjiang].

OBJECTIVE: We isolated myxobacteria in saline-alkaline soils of Xinjiang using a prey-predator strategy where the prey bacteria can induce the predator myxobacteria to form visible fruiting body, and evaluated intrinsic relationships between prey and predator myxobacteria. METHODS: Sixteen bacteria with inductive effects of fruiting body formation were obtained, and then used as preys to isolate the myxobacteria. RESULTS: A total of 55 myxobacteria strains were isolated from 25 soil samples, which were identified to the genera of Myxococcus, Corallococcus, Pyxidicoccus, Cystobacter, and Nannocystis. Besides, 6 unpurified isolates were believed to be myxobacteria. All the 16 prey bacteria had preferable inductive effects on Myxococcus spp., whereas Pyxidicoccus spp. and Cystobacter spp. were only induced by Gram-positive strains. CONCLUSION: The prey-predator strategy provided a new and more effective way to isolate myxobacteria.

[Isolation and identification of cultivable myxobacteria in the rhizosphere soils of medicinal plants].

OBJECTIVE: We isolated myxobacteria and explored their diversity from the rhizosphere soils of some medicinal plants. METHODS: We used the helper bacteria baiting technique to isolate myxobacteria from the rhizosphere soils collected in South China Botanical Garden and Nanling National Forest Park. The myxobacteria were identified by morphological characteristics and 16S rDNA gene sequences analysis. RESULTS: A total of 50 strains were isolated from 22 soil samples, which were identified into 7 genera, Myxococcus (18), Corallococcus (11), Cystobacter (7), Archangium (8), Stigmatella (1), Chondromyces (4) and Pyxidicoccus (1). The dominant genera were Myxococcus and Corallococcus. CONCLUSION: Environmental factors were associated with the diversity of myxobactria. Myxobacteria better adapt in high organic matter content and neutral pH environments. The strains of Myxococcus and Corallococcus had a good adaptability for different pH. Meanwhile, the dependence of the strains of Myxococcus and Cystobacter on organic carbon content was not too obvious, and they can also be found in the poor soils. Our findings provided an important scientific base for the development and utilization of myxobacteria resources.